Topoisomerases are enzymes which modify the topological state of DNA and have been shown to be involved in DNA replication and transcription. Since the process of retroviral replication, integration and expression are likely to require topological changes in either the integrated or unintegrated provirus, we have begun to determine if topoisomerases play a critical role in retrovirus growth or infection, or if topoisomerases might be a target for the control of retroviral infections. We had previously described a novel topoisomerase I (topo I) activity associated with various retrovirus and lentivirus particles, and showed that the specific topo I inhibitor, camptothecin (CPT), could block HIV infection of H9 cells in tissue culture. Using spleen focus-forming virus (SFFV) induction of splenomegaly as an in vivo model, we have shown that CPT can block SFFV-induced diseases when coinjected with the virus into susceptible mice. To determine the ability of CPT to affect chronic virus production by infected cells, we treated CF2Th (dog thymus) cells producing equine infectious anemia virus with CPT. Continuous exposure to low levels of the drug resulted in 85-92% inhibition of virus produc- tion, as measured by RT released into the culture media, and similar levels of reduction in viral protein production by immunofluorescence and radioimmunoprecipitation analysis.